Bacteria are widely used in life science, as source for food products, for vaccine components and for biotechnological production of recombinant proteins. For production-scale microbial fermentations, high cell densities are often desired and thus much research and effort has been put into optimization of culture conditions.
Generally, cell culture and microbial fermentation can be performed in batch, fed-batch, continuous or perfusion mode, or in combinations thereof.
Animal cells generally grow slowly, are fragile and necessitate gentle culture conditions. For animal cells, perfusion culture is the preferred mode. In perfusion mode, feed solutions are fed to a bioreactor continuously and spent medium is constantly removed. The advantage of perfusion culture over fed-batch culture is that if any inhibitory metabolites are generated during growth, these metabolites will continuously be removed and thus their growth inhibitory effect will be minimized. During perfusion culture, the majority of the animal cells is retained in the bioreactor. This is often achieved by filtration. In such set-ups, high tangential velocity generated by crossflow or spinning filters may be used to keep the filter surfaces clean. Also, methods based on alternating tangential flow (ATF) have been developed which cause animal cell aggregates to wash back into the vessel removing a potential blockage (see e.g. U.S. Pat. No. 6,544,424, Furey J (2002) Genetic Engineering News 22: 62-63 and WO2005095578).
Compared with animal cells, bacterial growth rates are generally much faster and necessitate higher feeding rate, and lead to faster accumulations of inhibitory products. Consequently, industrial scale fermentation of bacteria is mostly done in batch or fed-batch mode. Efforts to obtain higher yields have mainly focussed on the improvement of culture media including increased agitation rates and vigorous delivery of gases into the culture and controlled feeding strategies which are not applicable to animal cell culture, which requires more gentle conditions. Perfusion-like forms of culturing bacteria, such as dialysis culture, have been successful with a number of bacteria (Schultz and Gerhardt (1969) Microbiol. Mol. Biol. Rev 33:1), including Neisseria gonorrhoeae (Gerhardt and Heden (1960) Proc. Soc. Exp. Biol. Med. 105:49) but are generally difficult to scale-up. Jung and Lowitt ((2010), J Chem Technol Biotechnol, 85: 1250-1259) describe perfusion culture of lactic acid bacteria in a pilot scale membrane bioreactor. The growth of bacteria were restricted due to serious membrance fouling on the product membrance by extracellular lipopolysaccharides and glycoproteins.
Fuchs et al. (2002) J. Biotechnol. 93:243 describe a scaled-up dialysis fermentation method for Escherichia coli which results in increased cell densities and protein production. There remains a need for easy, robust, and upscalable methods for the fermentation of bacteria, in particular fastidious bacteria, which result in high cell densities and/or high production of biopolymers.